Histone covalent modifications are tightly linked to the regulation of eukaryotic gene expression. Monoubiquitylation of histone H2B (H2Bub) in the budding yeast, Saccharomyces cerevisiae, is associated with transcriptional activation, and regulates a novel trans-histone pathway that leads to methylation of histone H3 on lysines 4 (H3K4) and 79 (H3K79). The broad goals of the proposed research are to understand how H2Bub is regulated and how this histone modification influences transcription and stationary phase, a specialized quiescent state that results from nutrient deprivation. These goals will be pursued in three specific aims. In Specific Aim 1, the role of H2Bub in Pol II transcription will be defined using genetic and molecular approaches to determine if the histone modification co-operates with histone chaperones to regulate chromatin stability during Pol II initiation and elongation. In Specific Aim 2, the factors that regulate H2Bub will be studied by focusing on the roles of the E3 ligase, Bre1, in activation of the E2, Rad6. In Specific Aim 3, molecular and genetic approaches will be employed to identify the factors that regulate the deubiquitylation of H2Bub when cells enter stationary phase and the rapid ubiquitylation of H2B when cells re-enter the growth phase. The role of H2Bub in stationary phase will be investigated using molecular and cell biological approaches to determine whether H2Bub regulates the formation of quiescent cells. Aberrations in transcription can frequently result from epigenetic changes that are linked to the modification state of histones. These changes, in turn, have been associated with a number of disease states. The proposed studies on H2B ubiquitylation have relevance to cancer through the role of H2Bub in the trans- histone methylation of H3K4. The human SET domain protein, MLL1, is an H3K4 histone methyltransferase that is frequently rearranged in acute leukemias, and thus studies in yeast could provide insight into the roles of aberrant H3K4 methylation in tumor cells. In addition, studies on the role of H2Bub in cellular quiescence could provide a new understanding of aging and neurodegenerative diseases.